Hepatitis C virus diagnostics: technology, clinical applications and impacts.

Infection with hepatitis C virus (HCV) can cause several significant health problems. Sensitive and specific assays for antibodies to HCV are available to identify and confirm individuals infected with HCV. Evaluation of the clinical impact of HCV infection in a patient includes measuring liver biochemistries and possibly liver biopsy. Molecular assays such as HCV genotype identification and qualitative/quantitative HCV RNA analyses may be valuable in considerations of prognosis and therapy. Further refinement of antibody screening and confirmatory assays and standardization of molecular testing are necessary to optimize testing and thus fully characterize the diagnosis of HCV infection.

Use of a new vitamin C-deficient diet in a depletion/repletion clinical trial.

To conduct an inpatient study on the recommended dietary allowance (RDA) for vitamin C, we developed a unique vitamin C-deficient diet using a nutrient database and selective menus. Fourteen different menus were developed offering > 300 items with 0-2.4 mg vitamin C per serving. During the 4-6 mo volunteers were hospitalized, daily dietary vitamin C was restricted to < or = 5.0 mg. The mean daily dietary vitamin C intake was < 3.9 mg for the seven study subjects. With concurrent supplementation, the diet provided > or = 85% of the RDA for 17 essential nutrients. Within 3 wk of admission the diet induced vitamin C deficiency as indicated by plasma concentrations, which decreased from 23 +/- 6.9 to 6.9 +/- 2.0 mumol/L. Daily intake of vitamin C and five other nutrients was determined by nutrient database analyses. Mean energy, protein, the iron were 105-185% of the RDA and total and saturated fat were 32% and 10% of energy, respectively. Weight and nutritionally relevant indexes remained normal. Dietary adherence, calculated by the number of days with < or = 5.0 mg vitamin C per total study days, was 88-98% per repletion dose. Computer analyses of menu selections permitted individual preferences to be met while restricting vitamin C intake to < or = 5.0 mg/d. There were no complications from the diet during the depletion and repletion phase. With this diet, ascorbic acid pharmacokinetics for escalating doses could be determined in healthy volunteers.

MCV as a guide to phlebotomy therapy for hemochromatosis.

BACKGROUND: A multitude of recommendations exist for laboratory assays to monitor the pace and endpoints of phlebotomy therapy for hemochromatosis. All of these recommendations rely on an assessment of storage iron to guide treatment, and none have been prospectively evaluated. STUDY DESIGN AND METHODS: Nine consecutive patients underwent serial monitoring of Hb, MCV, transferrin saturation, and ferritin during weekly phlebotomy to deplete iron stores (induction therapy) and less frequent sessions to prevent iron reaccumulation (maintenance therapy). Changes in MCV and Hb were used to guide the pace of phlebotomy over a median of 7 years of follow-up. RESULTS: During induction therapy, the MCV increased transiently because of reticulocytosis and then stabilized for a prolonged period before decreasing more sharply, which reflected iron-limited erythropoiesis. Iron depletion was achieved after a median of 38 phlebotomies and removal of 9.0 g of iron. Maintenance phlebotomy was targeted to maintain the MCV at 5 to 10 percent below prephlebotomy values and the Hb at >13 g per dL. Transferrin saturation fluctuated considerably during treatment, but remained below 35 percent during MCV-guided maintenance therapy. Ferritin values were not useful guides to the pace of phlebotomy. The median maintenance therapy phlebotomy interval was 7.5 weeks (range, 6-16), which corresponded to an average daily iron removal of 35 to 67 microg per kg. Most patients showed evidence of iron reaccumulation at phlebotomy intervals of 8 weeks or more. CONCLUSION: The MCV is an inexpensive, precise, physiologic indicator of erythropoietic iron availability. When used in conjunction with the Hb, it is a clinically useful guide to the pace of phlebotomy therapy for hemochromatosis.

Pharmacokinetic model of ascorbic acid in healthy male volunteers during depletion and repletion.

PURPOSE: To develop a new pharmacokinetic model for ascorbic acid (vitamin C) since no previously published model describes ascorbic acid absorption and disposition over a broad physiologic range of doses and plasma concentrations. METHODS: A new model was developed through exploratory simulations. The model was fitted to pharmacokinetic data obtained from seven healthy volunteers who underwent ascorbic acid depletion then gradual repletion. Concentrations of ascorbic acid were measured in plasma and urine. Final pharmacokinetic model parameter estimates were obtained using nonlinear regression analysis. RESULTS: The new model included saturable absorption, distribution and renal tubular reabsorption parameters. The model described ascorbic acid concentrations in plasma, cells, and urine during depletion and gradual repletion phases with a residual error less than 15%. CONCLUSIONS: The model was useful for obtaining a new understanding of the likely causes for the complex concentration-time profile observed during gradual repletion. At doses of 200 to 2500 mg per day, the plateau in pre-dose concentrations is largely due to apparent saturation of tissue uptake and less a function of oral bioavailability and renal excretion than previously thought.

Hepatitis C in asymptomatic blood donors.

Among 248 asymptomatic blood donors positive for antibody to hepatitis C virus (anti-HCV) enrolled in a long-term prospective study, 86% had chronic HCV infection and 14% appeared to have recovered as assessed by serial determinations of serum alanine aminotransferase (ALT) levels and HCV RNA by polymerase chain reaction. Established parenteral risk factors for HCV transmission were identified in 75% of donors. In addition, there was a strong independent association between HCV positivity and cocaine snorting, suggesting that shared snorting devices may be a covert route of parenteral transmission. Ear piercing in males was also significantly associated with transmission. There was no evidence for sexual spread. Although the majority of HCV carriers had both biochemical and histological evidence of chronic viral hepatitis, the extent of liver injury was generally mild. Among a larger population of 280 HCV RNA-positive donors, 17% had repeatedly normal ALT levels, 45% had levels that did not exceed twice, and only 22% had levels that exceeded five times the upper limit of the normal range. Among 81 patients who underwent liver biopsy, only 13% had evidence of severe hepatitis (8%) or cirrhosis (5%), despite a duration of infection that generally exceeded 15 years. No severe histological lesions were observed in blood donors with chronic HCV infection who had repeatedly normal ALT levels. In both donors and blood recipients, the frequency of severe morbidity or mortality related to HCV infection was less than 10% during the first two decades of infection. Further long-term studies are required to see if the progression to severe outcomes continues to accrue at this slow pace or whether it accelerates during subsequent decades.

Routes of infection, viremia, and liver disease in blood donors found to have hepatitis C virus infection.

BACKGROUND: For many people infected with the hepatitis C virus (HCV), the route of exposure, risk of transmission, and severity of associated liver disease are unknown. We studied these variables in people who donated blood voluntarily. METHODS: Blood donors who tested positive for HCV antibodies on enzyme immunoassay were classified according to whether the results of a confirmatory second-generation recombinant immunoblot assay (RIBA) for HCV were positive, negative, or indeterminate. The evaluations also included an assessment of risk factors, a physical examination, serial determinations of alanine aminotransferase levels and HCV serologic assays, a polymerase-chain-reaction assay for HCV RNA, testing of sexual contacts and family members, and liver biopsies in some participants who were HCV-positive by RIBA. RESULTS: A total of 481 donors were studied, among whom 248 were positive for HCV by RIBA, 102 had indeterminate results, and 131 were HCV-negative. In a logistic-regression analysis, significant risk factors for HCV infection among the HCV-positive participants were a history of blood transfusion in 66 (27 percent; P < 0.001 for the comparison with RIBA-negative donors), intranasal cocaine use in 169 (68 percent, P < 0.001), intravenous drug use in 103 (42 percent, P = 0.001), sexual promiscuity in 132 (53 percent, P = 0.002), and ear piercing among men (P < 0.05). Nine of 85 sexual partners of HCV-positive donors were anti-HCV-positive; 8 had used intravenous drugs or received transfusions. HCV RNA was found in 213 HCV-positive donors (86 percent), 3 who had indeterminate results by RIBA (2 of these 3 tested positive with a more specific, third-generation RIBA), and none who were HCV-negative. Of the HCV-positive donors, 69 percent had biochemical evidence of chronic liver disease; among 77 donors positive for HCV by RIBA who underwent liver biopsy, 5 had severe chronic hepatitis or cirrhosis, 66 had mild-to-moderate chronic hepatitis, and 6 had no evidence of hepatitis. CONCLUSIONS: Among volunteer blood donors, prior blood transfusion, intranasal cocaine use, intravenous drug use, sexual promiscuity, and ear piercing in men are risk factors for HCV infection. The high frequency of intravenous drug use was unexpected, because these donors had denied such use when questioned directly at the time of their blood donations.

Stability of HCV-RNA level and its lack of correlation with disease severity in asymptomatic chronic hepatitis C virus carriers.

This study examines the relationship between HCV-RNA levels and disease severity in 60 individuals with chronic hepatitis C virus infection. HCV-RNA levels were quantified by the branched DNA (bDNA) assay in 445 samples (median: eight samples per patient) obtained over a median of 40.4 months (95% confidence interval (CI): 37.0-42.5). The median log HCV-RNA level was 6.77 (95% CI: 6.62-6.92) molecular equivalents/mL (MEQ/mL). The median log range of HCV-RNA levels in individual patients over the course of the study was 0.89 (95% CI: 0.69-1.16). HCV-RNA level varied over time by less than one log in 62% of patients, by 1-1.5 logs in 22% and by greater than 1.5 logs in only 17%. Univariate analysis, revealed an inverse association between HCV-RNA levels and ALT levels (P=0.037). Univariate and logistic regression analysis showed no significant association between HCV-RNA levels and either the degree of inflammation or fibrosis. In contrast, there was a significant positive association between alanine aminotransferase (ALT) levels and histological activity especially in individuals with ALTs> 100 IU/L. Hence, HCV-RNA levels: (i) almost always fell within the dynamic range of the bDNA assay; (ii) were stable in asymptomatic chronically infected patients, with only a small proportion of patients exceeding a range of 1.5 logs; (iii) did not correlate with either the extent of inflammation or degree of fibrosis. In contrast, there was a strong association between ALT level and the histological severity of liver disease.

Vitamin C pharmacokinetics in healthy volunteers: evidence for a recommended dietary allowance.

Determinants of the recommended dietary allowance (RDA) for vitamin C include the relationship between vitamin C dose and steady-state plasma concentration, bioavailability, urinary excretion, cell concentration, and potential adverse effects. Because current data are inadequate, an in-hospital depletion-repletion study was conducted. Seven healthy volunteers were hospitalized for 4-6 months and consumed a diet containing <5 mg of vitamin C daily. Steady-state plasma and tissue concentrations were determined at seven daily doses of vitamin C from 30 to 2500 mg. Vitamin C steady-state plasma concentrations as a function of dose displayed sigmoid kinetics. The steep portion of the curve occurred between the 30- and 100-mg daily dose, the current RDA of 60 mg daily was on the lower third of the curve, the first dose beyond the sigmoid portion of the curve was 200 mg daily, and complete plasma saturation occurred at 1000 mg daily. Neutrophils, monocytes, and lymphocytes saturated at 100 mg daily and contained concentrations at least 14-fold higher than plasma. Bioavailability was complete for 200 mg of vitamin C as a single dose. No vitamin C was excreted in urine of six of seven volunteers until the 100-mg dose. At single doses of 500 mg and higher, bioavailability declined and the absorbed amount was excreted. Oxalate and urate excretion were elevated at 1000 mg of vitamin C daily compared to lower doses. Based on these data and Institute of Medicine criteria, the current RDA of 60 mg daily should be increased to 200 mg daily, which can be obtained from fruits and vegetables. Safe doses of vitamin C are less than 1000 mg daily, and vitamin C daily doses above 400 mg have no evident value.

Volunteer blood donors with antibody to hepatitis C virus: clinical, biochemical, virologic, and histologic features. The Hepatitis C Study Group.

OBJECTIVE: To assess the clinical significance of antibody to hepatitis C virus (anti-HCV) in volunteer blood donors. DESIGN: Prospective cohort study. SETTING: National Institutes of Health Clinical Center, a tertiary referral research hospital. PATIENTS: 60 anti-HCV-positive blood donors, divided into three groups of 20 persons each: Group I had normal alanine aminotransferase levels, group II had levels elevated to values less than twice the normal range, and group III had levels elevated to values greater than twice the normal range. MEASUREMENTS: Medical history, results of laboratory and virologic testing, and percutaneous liver biopsy findings. RESULTS: Participants with normal alanine aminotransferase levels were older and more often female than those with abnormal levels. The source of infection, duration of disease, symptom score, and amount of alcohol consumed were similar in the three groups. Hepatitis C virus RNA was detectable in 85% of participants, more commonly in the groups with elevated alanine aminotransferase levels (95%) than in the group with normal levels (65%); however, titers were similar in all groups. Examination of liver biopsy specimens showed chronic hepatitis in 54 participants (90%) and cirrhosis in 1 participant. The only normal liver biopsy specimens (n = 3) were those from participants who were HCV RNA negative and had normal alanine aminotransferase levels. CONCLUSIONS: Most blood donors with anti-HCV have chronic hepatitis C regardless of their serum alanine aminotransferase levels. Donors with normal alanine aminotransferase levels and no HCV RNA in their serum generally have normal liver histologic findings or minimal changes and have probably recovered from HCV infection.

Analysis of hepatitis G virus (HGV) RNA, antibody to HGV envelope protein, and risk factors for blood donors coinfected with HGV and hepatitis C virus.

Serologic, biochemical, and molecular analyses were used to study hepatitis G virus (HGV), antibody to the HGV envelope protein (anti-E2), risk factors, clinical significance, and the impact of HGV on coexistent hepatitis C virus (HCV). Among 329 donors with confirmed HCV infection, 12% were HGV RNA-positive and 44% were anti-E2-positive (total exposure, 56%). HGV RNA and anti-E2 were mutually exclusive except in 9 donors (1.5%); 8 of 9 subsequently lost HGV RNA but anti-E2 persisted. HGV had little impact on alanine aminotransferase, aspartate aminotransferase, or gamma-glutamyl transpeptidase in donors with HGV infection alone or those coinfected with HCV. A multivariate analysis showed that intravenous drug abuse was the leading risk factor for HGV transmission, followed by blood transfusion, snorting cocaine, imprisonment, and a history of sexually transmitted diseases. In summary, HGV and HCV infections were frequently associated and shared common parenteral risk factors; HGV did not appear to cause hepatitis or to worsen the course of coexistent hepatitis C.